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* Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, and
Multidisciplinary Oncology Center, University Hospital, Lausanne, Switzerland;
Division of Oncology, Laboratory of Tumor Immunology, University Hospital, Geneva, Switzerland; and
Ludwig Institute for Cancer Research, Lausanne Branch, University of Lausanne, Epalinges, Switzerland
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The differentiation Ag Melan-A is selectively expressed by normal cells
of the melanocytic lineage including skin melanocytes and by the large
majority of malignant melanomas (10). HLA-A2-restricted,
Melan-A-specific CD8+ T cells primarily recognize
peptides 27–35 and 26–35 (11). The repertoire of
Melan-A-specific T cells detectable ex vivo by staining with
HLA-A2/Melan-A peptide multimers (referred to as multimers hereafter)
in HLA-A2-expressing individuals, including both normal donors and the
majority of melanoma patients, is unique, as it comprises
10-3 of circulating
CD8+ T cells phenotypically naive
(CD45RA+CCR7+)
(12, 13), being therefore the only known human Ag-specific
naive repertoire thus far accessible to analysis. We have recently
undertaken clinical trials of Melan-A26–35
peptide vaccination in HLA-A2 melanoma patients. In the course of the
vaccination, one patient developed a vigorous Ag-specific T cell
response as assessed by staining of PBL with HLA-A2/Melan-A peptide
multimers (14). In this work, we performed a longitudinal
analysis of this response both at the polyclonal and at the monoclonal
level in samples encompassing a time period of about 1 year after the
beginning of vaccination and compared those to both a preimmune
sample and a sample containing Ag-specific lymphocytes infiltrating an
autologous metastatic lesion.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Detailed clinical history and vaccination protocol for melanoma patient LAU 337 have previously been reported (14). The patient started being vaccinated in April 1999 and received four i.m. injections (at days 1, 28, 56, and 84) of synthetic peptides influenza Matrix58–66 (GILGFVFTL) and Melan-A26–35 (EAAGIGILTV) (100 µg each at each injection), in 600 µl of adjuvant SB-AS2 (provided by GlaxoSmithKline, Rixeusert, Belgium). Four additional injections of the same peptides, but administered s.c. in 600 µl Montanide adjuvant provided by SEPPIC (Paris, France), were given at days 147, 175, 204, and 301. From day 203 to 210 the patient received 5 x 106 IU of human (h)4rIL-2 (Proleukin; Roche, Reinach, Basel, Switzerland), twice a day. A s.c. metastasis of the cheek was surgically removed for esthetical reasons at day 29 from the beginning of vaccination. By the end of 1999 complete regression of some s.c. metastases and partial regression of one lung metastasis was observed, whereas other lesions remained stable. The patient died in May 2000 because of disease progression. Multimer+CD8+ T cells (see next paragraph) were purified from PBL ex vivo by flow cytometry cell sorting and expanded in vitro with PHA and allogeneic feeder cells as bulk cultures or cloned by limiting dilution as described (15). Metastatic tissue of the cheek was finely minced with needles in sterile culture medium. The cell suspension was then cultured in medium containing 100 U/ml hrIL-2 and 10 ng/ml hrIL-7 during 2 wk before analysis. Melan-A-specific CTL clones were derived from tumor-infiltrating lymphocytes (TIL) by limiting dilution cultures. All bulk cultures and clones were subsequently expanded by periodical (3–4 wk) restimulation into microtiter plates together with irradiated feeder cells in the presence of PHA, and hrIL-2. Ag recognition was assessed functionally by chromium release assay as previously described (16). The percentage of specific lysis was calculated as follows: 100 x [(experimental - spontaneous release)/(total - spontaneous release)].
A2/peptide multimers and flow cytometry immunofluorescence analysis
PE-conjugated multimeric HLA-A2/peptide complexes were
synthesized as described (6, 17). As the antigenic
peptide, the Melan-A26–35 A27L analog
(ELAGIGILTV), which has a higher binding affinity and stability than
parental peptide (EAAGIGILTV) (18), was used.
Interchangeability of parental Melan-A decapeptide and A27L analog in
terms of staining specificity has previously been assessed
(17). Cell samples were stained with multimers (4.5
µg/ml) in PBS 0.2% BSA during 1 h at room temperature, washed
once in the same buffer, stained with mAbs during 30 min at 4°C,
washed again, and analyzed by flow cytometry. Anti-CD8 and
anti-CD45RA mAbs were purchased from BD Biosciences (San Jose, CA).
For TCR 
-chain variable segment (BV) repertoire analysis a panel of
21 anti-BV mAbs (purchased from Immunotech, Marseilles, France) was
used. Staining was performed as previously described (19).
MHC-peptide/TCR dissociation experiments were performed as detailed
elsewhere (16) by using an excess of unlabeled multimers
to avoid rebinding of PE-labeled multimers after their dissociation
from the TCR. Intensity of multimer fluorescence at each time point
analyzed was expressed as the natural logarithm of the normalized
fluorescence at t0. Data analysis was
performed using CellQuest software (BD Biosciences).
CDR3 size analysis of TCR BV transcripts, sequencing of PCR products, and quantitative assessment of selected Melan-A-specific TCR clonotypes
The complementarity-determining region (CDR)3 of the
PCR-amplified TCR BV1–24 transcripts was analyzed using a run-off
procedure as described previously (20, 21). The run-off
products were then run on an automated sequencer in the presence of
fluorescent size markers. The length of the DNA fragments and the
fluorescence intensity of the bands were analyzed with Base ImagIR
software (LI-COR Biotechnology Division, Bad Homburg, Germany). TCR
BV-BC PCR products derived from sorted populations were cloned with
TOPO TA cloning kit (Invitrogen, Carlsbad, CA). One Shot TOP10
chemically competent Escherichia coli (Invitrogen) were
transformed and plated for blue/white color selection on medium
containing
5-bromo-4-chloro-3-indolyl--D-galactopyranoside.
Plasmid DNA was extracted from white colonies using the Qiagen Plasmid
Mini kit (Qiagen, Hilden, Germany) and sequenced using Thermo Sequenase
fluorescent-labeled primer cycle sequencing kit with 7-deaza-dGTP
(Amersham Pharmacia Biotech, Little Chalfont, U.K.). Quantification of
selected T cell clones was performed as previously described (21, 22).
The sequences of clonotypic primers were as follows: BV3, 5'-GTGCCCAACCCGAGAAAACT-3'; BV5, 5'-GCTGATTGCTGTCCAAGCTG-3'; BV17, 5'-CTGCTCATTGCCAGGTATAC-3'; BV14, clone 2C2 5'-CCCGGTGCCAGATAAACTGCT-3', clone 1D3, 5'-GGTTGTCCCCGATAAACTGCT-3', clone 4E3, 5'-AACCCAGGAGCCCTCCCAAACT-3', and clone 4C8, 5'-TTCAGTCCGTCGATAGGGCT-3'.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The clinical course and vaccination protocol for melanoma patient
LAU 337 are detailed in Materials and Methods. Blood samples
were obtained before peptide vaccination (day -5) and after two (day
39), four (day 100), six (day 176), and seven (days 211 and 300)
injections. The frequency of Melan-A-specific T cells in pre- and
postimmune peripheral blood samples was assessed by costaining with
anti-CD8 mAb and multimers (17). Before peptide
vaccination (day -5), we detected a relatively high frequency of
multimer+ T cells (0.1% of total
CD8+ T cells; Fig. 1
) that exhibited a mixed phenotype
composed of both CD45RAbright and
CD45RAlow cells. We have previously observed this
mixed phenotype in multimer+ T cells from 30%
of HLA-A2 melanoma patients, whereas in the remaining patients (e.g.,
LAU 203) and in all normal donors analyzed (e.g., HD 604), the majority
of multimer+ T cells were
CD45RAbright (12). The percentage of
circulating multimer+ T lymphocytes significantly
increased already at day 39 after the beginning of vaccination (0.7%
of total CD8+ T cells) and accounted for 2.2,
1.8, 1, and 2.3% of total CD8+ T cells at days
100, 176, 211, and 300 postvaccination, respectively. It is of note
that the increase was mostly confined to the
multimer+CD45RAlow T cell
compartment (Fig. 1).
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for the line corresponding to day 211
postimmune and summarized in Table I,
lines from postimmune samples recognized Melan-A peptides with
higher avidity as compared with the preimmune line. At this time point
as well as at day 176, but not at day 39, the hierarchy of peptide
Melan-A nona- vsdecapeptide recognition was inverted in the postimmune
as compared with the preimmune line (Fig. 2A). Increased
functional avidity of Ag recognition resulted in improved lysis of
Melan-A-expressing tumor cells (Fig. 2B and Table I). It is
of note that, although all the populations tested were stained by
multimers with roughly comparable efficiency (Fig. 2C), the
TCR complexes formed with the lower-avidity population were less stable
than the ones formed with the high-avidity population, as observed
following the kinetics of multimer staining decay (Fig. 2D).
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Due to the high frequency of
CD8+multimer+ T cells in
postimmune samples, their BV usage could be assessed ex vivo by
combining staining with anti-CD8 mAb, multimers, and a panel of
mAbs directed against single BV subfamilies as previously described
(19). Several
CD8+multimer+ T cell
subpopulations, each expressing a distinct BV, were detected at each
time point analyzed. However, five subpopulations (expressing BV1, BV3,
BV5, BV14, or BV17) clearly dominated the response all along the
vaccination period. Indeed, the sum of multimer+
T cells expressing these BV regions accounted for 75–85% of the total
multimer+ T cell population at different time
points of the analysis (Fig. 3A). The relative proportion
of each subpopulation moderately (e.g., BV17) or significantly (e.g.,
BV3, BV5.1) varied among the different time points analyzed. To further
analyze the clonal composition of the multimer+ T
cell populations along the vaccination period,
multimer+ T cells were isolated ex vivo from
postimmune blood samples by multimer-guided cell sorting and directly
submitted to analysis of TCR -chain V segment and CDR3 length by
spectratyping (23). In contrast to the bell-shaped pattern
characteristic of polyclonal T cell populations that was obtained for
multimer negative fractions (for an example, see Fig. 3B,
bottom line), CDR3 size profiles obtained for
multimer+ populations displayed prominent peaks
that indicated the accumulation of recurrent size transcripts. For each
BV a unique peak (e.g., BV3) or multiple peaks (e.g., BV14) were
detected. Prominent peaks were detected mostly in the case of dominant
populations but also for some of the nondominant ones (e.g., BV2, BV13,
and BV16), indicating that clonal expansions, albeit of lower
amplitude, could also be present in those latter. Of note, in several
instances (e.g., BV3) prominent peaks as well as their CDR3 size
profiles were common between some or even all samples analyzed,
suggesting the presence of Melan-A-specific T cell clonotypes
persisting along the immune response.
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To analyze the Melan-A-specific response at the clonal level,
monoclonal populations were derived from samples corresponding to days
176 and 211 after vaccination, by cloning the sorted
CD8+multimer+ T cell
fractions under limiting dilution conditions as previously described
(24). Sequences of BV, CDR3, and junctional regions of the
isolated clones (grouped according to the BV) are reported in Table II. The analysis of the BV used by the
isolated clones (a total of 17) confirmed that clones bearing the BV3,
BV5.1, BV14, and BV17 were among the most frequently used by
multimer+ T cells. Somewhat surprisingly, no
BV1-using clones were isolated. Whereas in the case of BV3 (four clones
isolated), BV5 (four clones isolated), and BV17 (two clones isolated) a
single clonotype was found, four different clonotypes were identified
in the case of BV14. Avidity and fine specificity of Ag recognition of
single clonal populations was assessed in a functional chromium release
assay in the presence of graded concentrations of either
Melan-A27–35 or
Melan-A26–35 peptides as illustrated in Fig. 4A. In addition, the ability
of the clones to specifically lyse Melan-A-expressing tumors was also
assessed. Avidity of Ag recognition and tumor reactivity were roughly
similar for all isolated clones, irrespective of their BV usage and
comparable to that of the corresponding polyclonal monospecific
postimmune line (Fig. 4A and data not shown). For internal
comparison, clonal populations were also derived from limiting dilution
cloning of the preimmune polyclonal monospecific line. Eleven of the 14
Melan-A peptide-reactive clones used BV2 and recognized
Melan-A27–35 much more efficiently than
Melan-A26–35, and the remaining three used
BV13.1 and recognized Melan-A27–35 slightly
better than Melan-A26–35. Functional data from
one representative clone from each group are shown in Fig. 4B. Consistent with the data obtained on polyclonal
monospecific lines, these clones displayed a decreased level of tumor
reactivity as compared with the ones derived from postimmune samples
(Fig. 4, A and B, and data not shown).
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The frequency of individual Melan-A-specific T cell
clonotypes among the total multimer-sorted population in the different
postimmune samples was assessed using the previously described approach
with clonotypic primers pairing unique CDR3 (23). Seven
clonotypic primers corresponding to distinct CDR3 sequences in Table II
were synthesized and used to quantify the corresponding clonotypes
within the total BV-corresponding transcripts. Considering the
proportion of T cells expressing that particular BV segment in the
multimer+ fraction (Fig. 3A), the
proportion of the clone within multimer+ T cells
was then extrapolated. Clone 1B7 represented a high proportion (from 32
to 87%) of total BV3 transcripts in all postimmune samples (Fig. 5A). This individual clone was
present at a relatively low frequency (<0.1% of total
multimer+ cells) at day 39, whereas, at the
maximum of its expansion (day 176), it represented up to 11.3% (Fig. 5B) of the total multimer+ population,
corresponding to an absolute number of 106
cells per liter of blood. The same procedure was used to quantify the
remaining clonotypes. Clones 2C2, 4E3, and 4C8 represented, at the
maximum of their expansion, 12–15% of total BV14 transcripts each,
i.e., 3.3, 2.7, and 3.4% of multimer+ T cells,
respectively. The remaining three clones (6C3, 5B3, and 1D3) were
quantitatively minor, detectable only at days 176, 211, and 300, and
accounting at most for only 0.2–0.56% of the total
multimer+ T cell population (data not shown).
Thus, despite the relatively high number of clones isolated and due to
the complexity of the populations under study, this analysis only
covered 15% of cells composing the multimer+
population. It is of note that the different clones did not expand
simultaneously. Indeed, the proportion of clone 1B7, for example,
significantly increased during vaccination, peaking at day 176, and
declined thereafter, whereas a significant increase in the proportion
of clones 4C8 and 4E3 was measured only at day 300 (Fig. 5B). Thus, whereas some clones progressively disappeared
from the circulation after initial expansion, other clones initiated
their expansion only late after the beginning of vaccination. It is of
note that, using clonotypic primers, we failed to detect the clonotypes
isolated from the postimmune samples among both naive and
Ag-experienced specific cells in the preimmune sample (data not shown),
indicating that, before immunization, those clonotypes would be present
in both populations at a frequency <1/1000 cells. This observation is
consistent with previous data obtained in a murine model, indicating
that the frequency of clones expanded by vaccination was extremely low
in the preimmune repertoire (25).
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A).
Multimer+ T cells in TIL were isolated by sorting
and analyzed by spectratyping. Interestingly, the CDR3 size of some
prominent peaks was similar in TIL and in PBL from the closest date
(day 39), suggesting the presence of identical clonotypes (Fig. 6B). Indeed, as assessed by analysis with clonotypic primers
(Fig. 6C), two of the seven clones isolated from the PBL
were present in this metastasis. Clone 5B3, which was present at a very
low proportion in the PBL, represented 12% of TIL
multimer+ cells, whereas clone 1B7, which was
prominent among PBL (Fig. 5A), was quantitatively minor in
TIL. However, quantitative considerations should be taken with caution,
as in vitro culture of TIL could have significantly modified the
relative proportion of the clones. Six T cell clones were derived from
the TIL multimer+ population. As partially shown
in Fig. 4C for two representative clones, Melan-A-specific
clones derived from TIL displayed relatively high avidity of Ag
recognition and high tumor reactivity. Two clones used BV17. Analysis
of the CDR3 revealed that one of those was identical to clone 5B3. The
remaining four clones used BV20. In good agreement with this data as
well as with the results of spectratyping analysis, costaining of TIL
with multimers and anti-BV20 mAb showed that 21% of
multimer+ T cells in TIL used BV20 (Fig. 6A).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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60-fold, Table I) and did
not further increase afterward. Interestingly, improved tumor
recognition followed a similar kinetics, suggesting that, as previously
indicated by mass spectrometric analysis of naturally processed Melan-A
peptides (28), Melan-A27–35 could
be the main peptide species presented on melanoma cells. In contrast,
improved recognition of peptide Melan-A26–35
(the immunogen) became apparent only at a later time point (day 176)
and further increased at day 211, resulting in a >600-fold improved
recognition as compared with the preimmune sample. As a consequence of
these changes, the hierarchy of recognition of nona- vs decapeptide was
inverted in the later postimmune as compared with the preimmune sample.
Similar results, in terms of both avidity of Ag recognition as well as
relative efficiency of tumor recognition, were obtained with
representative clonal populations (Fig. 4). Second, BV chain usage in
preimmune CTL clones (BV2 and BV13.1) was different from that of the
dominant populations in postimmune samples. Clonal populations derived
from the latter exhibited a broadly comparable activity, regardless of
their BV usage. Together, these data favor a view of T cell repertoire
selection based on Ag-driven selection of clones able to recognize the
immunogen with high avidity. However, no obvious structural basis for
this selection was apparent, as no TCR BV and/or CDR3 sequence
constraints or homologies among the selected clones were identified
(Table II), in agreement with our previous finding that the repertoire
of Melan-A-specific T cells naturally selected at the tumor site was
diverse and mostly nonoverlapping among different individuals
(19, 23). According to the kinetic model of T cell signaling (29, 30, 31) the biological consequences of the TCR interaction with MHC/peptide complexes is primarily determined by its duration: only an optimal time of interaction would allow the completion of the multiple steps implicated in full T cell activation, eventually leading to T cell proliferation and clonal expansion. Results in mouse models of CD8+ T cell response to infectious diseases have indeed indicated that specific T cells derived from mice responding to primary infection were characterized by faster TCR/multimer dissociation rates than cells derived from mice responding to recall infection (32, 33). Similarly, in this study, polyclonal monospecific cells from postimmune samples displayed a mean dissociation kinetics significantly slower than that displayed by preimmune cells. However, in contrast to other reports (34), we did not detect significant differences in the multimer staining intensity of these populations. Consistent with these data, by using monoclonal CD8+ T cell populations specific for three tumor Ag-derived peptides including Melan-A, we have observed that the efficiency of staining with the corresponding multimers can considerably vary with staining conditions and does not always correlate with functional avidity of Ag recognition. In contrast, in each case we found a direct correlation between the latter and the stability of the corresponding TCR/multimer complexes (16). These data support the view that maturation of functional avidity during immune responses to cancer would result from the selective clonal expansion of T cells expressing TCR with relatively low dissociation rates from MHC/peptide complexes.
Longitudinal analysis of circulating Melan-A multimer+ populations along the vaccination period revealed that after an initial progressive increase in the frequency of detectable circulating multimer+ cells, this population remained quantitatively stable over time, representing between 1.1 and 2.3% of total circulating CD8+ T lymphocytes. However, the frequency of individual clones within this population considerably and asynchronously varied over time, with some clones declining after initial expansion and others expanding only at late time points. This could be the consequence of either different time of recruitment of multiple T cell clones, each with relatively limited growing potential and life span, and/or of differential trafficking of the latter from and/or to vaccination/tumor sites. Interestingly, in contrast with our results, similar studies of immune responses to SIV- and HIV-derived epitopes have shown that the frequency of some dominant clonal populations can be stable over relatively long periods of time (35, 36). An explanation for this difference could be related to the nature of the immunogen that elicits these CD8+ T cell responses. Indeed, whereas in the case of immune responses to intact pathogens or derived proteins a specific CD4+ Th cell component (which plays a central role in sustaining CD8+ T cell responses) (37) is most likely to be involved, this would in general not be the case upon immunization with short peptides corresponding to well-defined CD8+ T cell epitopes. In this case, only repeated peptide injections would be able to continuously recruit additional specific clonal populations. These observations might provide a rationale for the design of more efficient vaccination schedules.
It is of note that whereas patient LAU 337 and, to a much lesser
extent, some other melanoma patients enrolled in the same trial of
peptide vaccination mounted a Melan-A-specific response (mostly
confined to the CD45RAlow population)
concomitantly with peptide vaccination, other patients failed to do so
48 . The basis for the heterogeneity of responsiveness to
vaccination with Melan-A peptide is not clear at this stage but could
be related to the functional state of T cells before vaccination. In
this regard, a multimer+ population with an
Ag-experienced phenotype accounting for 50% of the total population
was already present in the preimmune sample of patient LAU 337,
indicating that in this patient a certain level of activation of
Melan-A-specific cells had already occurred spontaneously, most likely
in response to the autologous tumor. Further studies directly comparing
the T cell repertoire of naive vs Ag experienced Melan-A
multimer-positive T cells in responder patients could, in the future,
provide more insight into these questions.
Although the vigorous response to peptide vaccination of patient
LAU 337 was accompanied by transient regression of some metastases,
after 1 year of stable disease a rapid tumor progression occurred. One
possible explanation for the apparent dissociation between the observed
immunological and clinical responses is that the induced response,
although substantial, may be quantitatively insufficient. In humans,
the information on the potential levels of tumor-specific
CD8+ T cells which may correlate with clinical
responses is complex, often contradictory, and difficult to evaluate.
Vigorous specific responses accounting for as much as 3–4% of
circulating CD8+ T lymphocytes have been detected
following vaccination with a gp100 peptide analog, often in the absence
of significant tumor regression (5). In contrast, in other
studies, repeated long-term administration of melanoma Ag-derived
peptides induced clinical response in the absence of detectable
circulating specific T cells (3). High levels of specific
CD8+ T cells are detected in immune response to
viral infections. For example, as many as 44% of
CD8+ T cells specific for an immunodominant
EBV-derived epitope can be detected during the acute phase of the viral
infection (38). However, a level of 2% of specific
CD8+ T lymphocytes is apparently sufficient to
prevent disease progression in individuals infected with HIV
(39). Thus, taking into account the limited information
presently available, it is difficult to evaluate whether the level of
specific CD8+ T cells elicited in patient LAU 337
was quantitatively adequate for controlling tumor growth.
Interestingly, some of the clonotypes present in the circulation were
retrieved among lymphocytes infiltrating a s.c. tumor metastasis, thus
making it unlikely that tumor progression could be due to the inherent
inability of tumor-specific circulating cells to reach their target at
the tumor site. Alternative explanations for tumor escape could be
related to the loss of expression of either tumor Ags or Ag-presenting
molecules (40). Indeed, small areas of the s.c. metastasis
analyzed in this study showed a clear loss of expression of total MHC
class I molecules, although the large majority of the tumor tissue
analyzed showed normal MHC class I expression and uniformly expressed
Melan-A. Clearly, in the absence of a global assessment of MHC class I
and tumor Ag expression of massive progressive tumor metastases, the
impact of this element is difficult to evaluate. However, it is
intuitive that vaccination should be started earlier in the course of
diseases, when selective or global down-regulation of HLA and tumor Ag
expression is likely to be still modest.
It has been previously suggested that tumor-specific T cells present in the circulation could be, at least in some cases, anergic (41), or that Ag-specific T cells visualized with multimers may include populations displaying low avidity of Ag recognition and therefore not relevant for tumor recognition in physiological conditions (42). Moreover, persistent exposure of T cells to Ags could, under certain circumstances, result in various types of Ag-specific T cell dysfunction (43, 44, 45, 46). In a previous study (14) and in this study we have examined the functional state of the Melan-A multimer+ T cells in patient LAU 337. Consistent with a recent report in the case of patients responding to vaccination with the gp100-derived peptide analog (9), we failed to detect any sign of dysfunction in the postimmune Melan-A-specific population either after in vitro stimulation or ex vivo. However, it is of note that Melan-A multimer+ T cells elicited upon vaccination presented the phenotypical features of Ag-experienced T cells and not of effector T cells (47). Consistent with their phenotype these cells were able to readily secrete IL-2 upon short stimulation (3–6 h) with Ag. However, they failed to display significant lytic activity ex vivo but they developed the capacity to efficiently lyse Ag-expressing target cells upon in vitro expansion following stimulation with either mitogen or Ag (14). Thus, although the relative in vivo effectiveness of CD8+ T cells at these discrete differentiation stages for the control of tumor growth is presently unknown, the design of vaccination strategies aimed at inducing fully differentiated effector CTLs should be attempted. Finding the missing link between elicitation of tumor Ag-specific T cell responses and induction of efficient and stable tumor immune surveillance will most likely be the focus of tumor immunology in the next few years.
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Acknowledgments |
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Footnotes |
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2 D.V. and V.D. contributed equally to this work.
3 Address correspondence and reprint requests to Dr. Danila Valmori, Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Hôpital Orthopédique, Niveau 5, Aile Est, Avenue Pierre-Decker, 4-CH 1005 Lausanne, Switzerland. E-mail address: danila.valmori{at}inst.hospvd.ch
4 Abbreviations used in this paper: h, human; TIL, tumor-infiltrating lymphocyte; CDR, complementarity-determining region; BV, TCR -chain variable segment.
Received for publication December 21, 2001. Accepted for publication February 20, 2002.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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secretion by circulating CD8 T lymphocytes: implications of a novel approach for T cell monitoring in infectious and malignant diseases. J. Immunol. 166:7634.-chain usage reveals a large repertoire of Melan-A-specific CD8+ T cells in melanoma patients. J. Immunol. 165:533.This article has been cited by other articles:
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  G. Bioley, C. Dousset, A. Yeh, B. Dupont, N. Bhardwaj, G. Mears, L. J. Old, M. Ayyoub, and D. Valmori Vaccination with Recombinant NY-ESO-1 Protein Elicits Immunodominant HLA-DR52b-restricted CD4+ T Cell Responses with a Conserved T Cell Receptor Repertoire Clin. Cancer Res., July 1, 2009; 15(13): 4467 - 4474. [Abstract] [Full Text] [PDF]
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 L. Derre, M. Bruyninx, P. Baumgaertner, M. Ferber, D. Schmid, A. Leimgruber, V. Zoete, P. Romero, O. Michielin, D. E. Speiser, et al. Distinct sets of {alpha}{beta} TCRs confer similar recognition of tumor antigen NY-ESO-1157-165 by interacting with its central Met/Trp residues PNAS, September 30, 2008; 105(39): 15010 - 15015. [Abstract] [Full Text] [PDF]
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 L. Derre, M. Ferber, C. Touvrey, E. Devevre, V. Zoete, A. Leimgruber, P. Romero, O. Michielin, F. Levy, and D. E. Speiser A Novel Population of Human Melanoma-Specific CD8 T Cells Recognizes Melan-AMART-1 Immunodominant Nonapeptide but Not the Corresponding Decapeptide J. Immunol., December 1, 2007; 179(11): 7635 - 7645. [Abstract] [Full Text] [PDF]
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D. E. Speiser, P. Baumgaertner, C. Barbey, V. Rubio-Godoy, A. Moulin, P. Corthesy, E. Devevre, P.-Y. Dietrich, D. Rimoldi, D. Lienard, et al. A Novel Approach to Characterize Clonality and Differentiation of Human Melanoma-Specific T Cell Responses: Spontaneous Priming and Efficient Boosting by Vaccination J. Immunol., July 15, 2006; 177(2): 1338 - 1348. [Abstract] [Full Text] [PDF]
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M. J. Pittet, A. Gati, F.-A. Le Gal, G. Bioley, P. Guillaume, M. de Smedt, J. Plum, D. E. Speiser, J.-C. Cerottini, P.-Y. Dietrich, et al. Ex Vivo Characterization of Allo-MHC-Restricted T Cells Specific for a Single MHC-Peptide Complex J. Immunol., February 15, 2006; 176(4): 2330 - 2336. [Abstract] [Full Text] [PDF]
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 I. J. M. de Vries, M. R. Bernsen, W. J. Lesterhuis, N. M. Scharenborg, S. P. Strijk, M.-J. P. Gerritsen, D. J. Ruiter, C. G. Figdor, C. J.A. Punt, and G. J. Adema Immunomonitoring Tumor-Specific T Cells in Delayed-Type Hypersensitivity Skin Biopsies After Dendritic Cell Vaccination Correlates With Clinical Outcome J. Clin. Oncol., August 20, 2005; 23(24): 5779 - 5787. [Abstract] [Full Text] [PDF]
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K. Kedzierska, N. L. La Gruta, M. P. Davenport, S. J. Turner, and P. C. Doherty Contribution of T cell receptor affinity to overall avidity for virus-specific CD8+ T cell responses PNAS, August 9, 2005; 102(32): 11432 - 11437. [Abstract] [Full Text] [PDF]
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S. Yang, K.-Y. Tsang, and J. Schlom Induction of Higher-Avidity Human CTLs by Vector-Mediated Enhanced Costimulation of Antigen-Presenting Cells Clin. Cancer Res., August 1, 2005; 11(15): 5603 - 5615. [Abstract] [Full Text] [PDF]
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 H. Reijonen, R. Mallone, A.-K. Heninger, E. M. Laughlin, S. A. Kochik, B. Falk, W. W. Kwok, C. Greenbaum, and G. T. Nepom GAD65-Specific CD4+ T-Cells with High Antigen Avidity Are Prevalent in Peripheral Blood of Patients With Type 1 Diabetes Diabetes, August 1, 2004; 53(8): 1987 - 1994. [Abstract] [Full Text] [PDF]
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M. van Oijen, A. Bins, S. Elias, J. Sein, P. Weder, G. de Gast, H. Mallo, M. Gallee, H. van Tinteren, T. Schumacher, et al. On the Role of Melanoma-Specific CD8+ T-Cell Immunity in Disease Progression of Advanced-Stage Melanoma Patients Clin. Cancer Res., July 15, 2004; 10(14): 4754 - 4760. [Abstract] [Full Text] [PDF]
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 A. Zippelius, P. Batard, V. Rubio-Godoy, G. Bioley, D. Lienard, F. Lejeune, D. Rimoldi, P. Guillaume, N. Meidenbauer, A. Mackensen, et al. Effector Function of Human Tumor-Specific CD8 T Cells in Melanoma Lesions: A State of Local Functional Tolerance Cancer Res., April 15, 2004; 64(8): 2865 - 2873. [Abstract] [Full Text] [PDF]
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E. M.-L. Choi, J.-L. Chen, L. Wooldridge, M. Salio, A. Lissina, N. Lissin, I. F. Hermans, J. D. Silk, F. Mirza, M. J. Palmowski, et al. High Avidity Antigen-Specific CTL Identified by CD8-Independent Tetramer Staining J. Immunol., November 15, 2003; 171(10): 5116 - 5123. [Abstract] [Full Text] [PDF]
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I. J. M. de Vries, W. J. Lesterhuis, N. M. Scharenborg, L. P. H. Engelen, D. J. Ruiter, M.-J. P. Gerritsen, S. Croockewit, C. M. Britten, R. Torensma, G. J. Adema, et al. Maturation of Dendritic Cells Is a Prerequisite for Inducing Immune Responses in Advanced Melanoma Patients Clin. Cancer Res., November 1, 2003; 9(14): 5091 - 5100. [Abstract] [Full Text] [PDF]
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M. Ayyoub, D. Rimoldi, P. Guillaume, P. Romero, J.-C. Cerottini, D. Valmori, and D. Speiser Tumor-reactive, SSX-2-specific CD8+ T Cells Are Selectively Expanded during Immune Responses to Antigen-expressing Tumors in Melanoma Patients Cancer Res., September 1, 2003; 63(17): 5601 - 5606. [Abstract] [Full Text] [PDF]
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P.-Y. Dietrich, F.-A. Le Gal, V. Dutoit, M. J. Pittet, L. Trautman, A. Zippelius, I. Cognet, V. Widmer, P. R. Walker, O. Michielin, et al. Prevalent Role of TCR {alpha}-Chain in the Selection of the Preimmune Repertoire Specific for a Human Tumor-Associated Self-Antigen J. Immunol., May 15, 2003; 170(10): 5103 - 5109. [Abstract] [Full Text] [PDF]
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S. Oh, J. W. Hodge, J. D. Ahlers, D. S. Burke, J. Schlom, and J. A. Berzofsky Selective Induction of High Avidity CTL by Altering the Balance of Signals from APC J. Immunol., March 1, 2003; 170(5): 2523 - 2530. [Abstract] [Full Text] [PDF]
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V. Rubio-Godoy, V. Dutoit, Y. Zhao, R. Simon, P. Guillaume, R. Houghten, P. Romero, J.-C. Cerottini, C. Pinilla, and D. Valmori Positional Scanning-Synthetic Peptide Library-Based Analysis of Self- and Pathogen-Derived Peptide Cross-Reactivity with Tumor-Reactive Melan-A-Specific CTL J. Immunol., November 15, 2002; 169(10): 5696 - 5707. [Abstract] [Full Text] [PDF]
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S. Mandruzzato, E. Rossi, F. Bernardi, V. Tosello, B. Macino, G. Basso, V. Chiarion-Sileni, C. R. Rossi, C. Montesco, and P. Zanovello Large and Dissimilar Repertoire of Melan-A/MART-1-Specific CTL in Metastatic Lesions and Blood of a Melanoma Patient J. Immunol., October 1, 2002; 169(7): 4017 - 4024. [Abstract] [Full Text] [PDF]
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 V. Dutoit, V. Rubio-Godoy, M. J. Pittet, A. Zippelius, P.-Y. Dietrich, F. A. Legal, P. Guillaume, P. Romero, J.-C. Cerottini, R. A. Houghten, et al. Degeneracy of Antigen Recognition as the Molecular Basis for the High Frequency of Naive A2/Melan-A Peptide Multimer+ CD8+ T Cells in Humans J. Exp. Med., July 15, 2002; 196(2): 207 - 216. [Abstract] [Full Text] [PDF]
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H. Echchakir, G. Dorothee, I. Vergnon, J. Menez, S. Chouaib, and F. Mami-Chouaib Cytotoxic T lymphocytes directed against a tumor-specific mutated antigen display similar HLA tetramer binding but distinct functional avidity and tissue distribution PNAS, July 9, 2002; 99(14): 9358 - 9363. [Abstract] [Full Text] [PDF]
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